Lolium multiflorum (Italian ryegrass) has recently demonstrated itself to be poorly controlled with glyphosate in cereal crops of South Chile. The concentration of glyphosate necessary to reduce shoot length by 50% (ED50) in seedlings, after eight days of root contact was 7.3-fold in the resistant Vil-1 than in the susceptible (S) biotype. The obtained spray retention values were higher on S than the resistant (Vil-1) biotype. Contact angles measured on the adaxial surface of S and Vil-1 were similar. However, on the abaxial surface contact angles were of 63 degrees on Vil-1 as compared to 42 degrees on S. A greater glyphosate uptake was observed through the abaxial surface of S. Regarding translocation, glyphosate accumulated mainly in the tip of the treated leaf of Vil-1, 24 h after treatment. It was afterwards also well distributed to the rest of the leaves and roots, as in the susceptible biotype. Nevertheless, 14C-glyphosate remained higher in the foliar apex of Vil-1. Hence, resistance to glyphosate by the Vil-1 Lolium multflorum biotype seems to involve a lower uptake through the abaxial leaf surface and a different migration pattern.